In vitro biosynthesis of acetan using electroporated Acetobacter xylinum cells as enzyme preparations

Acetobacter xylinum strain NRRL B42 and its derivative RCGr1 produce a complex exopolysaccharide, acetan, containing glucose, mannose, glucuronic acid and rhamnose in a 4:1:1:1 molar ratio. The in vitro synthesis of acetan, employing electroporated cells as the enzyme system and the respective 14C-labelled sugar nucleotide precursors, is described. The synthesis of the prenyl-linked heptasaccharide repeat unit, already observed in EDTA-treated cells, was confirmed, as well as the formation of other saccharides not related to acetan biosynthesis, including a high molecular mass glucan. The acetan formed was characterized by gel filtration, specific radioactive labelling with each precursor and permethylation analysis. It was also shown that acetan contains acetyl residues and that using [14C]acetyl CoA as donor, radioactivity was detected both at the polysaccharide and at the prenyl-linked oligosaccharide stage.